Hardy S J, Holmgren J, Johansson S, Sanchez J, Hirst T R
Department of Biology, University of York, Great Britain.
Proc Natl Acad Sci U S A. 1988 Oct;85(19):7109-13. doi: 10.1073/pnas.85.19.7109.
In this paper we study the assembly, in vivo and in vitro, of a family of hexameric, heat-labile enterotoxins produced by diarrheagenic bacteria. The toxins, which consist of an A subunit and five B subunits, are assembled by a highly coordinated process that ensures secretion of the holotoxin complex. We show that (i) oxidation of cysteine residues in the B subunits is a prerequisite step for in vivo formation of B-subunit pentamers, (ii) reduction of dissociated B subunits in vitro abolishes their ability to reassemble, (iii) the kinetics of B-pentamer assembly in vivo can be mimicked under defined conditions in vitro, (iv) A subunits cannot associate with fully assembled B pentamers in vitro, and (v) A subunits cause an approximately 3-fold acceleration in the rate of B-subunit pentamerization in vivo, implying that A subunits play a coordinating role in the pathway of holotoxin assembly. The last finding is likely to be of general significance, since it provides a mechanism for preferentially excluding or favoring certain intermediates in the assembly of multisubunit proteins.
在本文中，我们研究了致泻性细菌产生的一类六聚体、热不稳定肠毒素在体内和体外的组装过程。这些毒素由一个A亚基和五个B亚基组成，通过一个高度协调的过程进行组装，该过程确保了全毒素复合物的分泌。我们发现：（i）B亚基中半胱氨酸残基的氧化是体内形成B亚基五聚体的先决步骤；（ii）体外解离的B亚基还原后丧失了重新组装的能力；（iii）在特定体外条件下可以模拟体内B五聚体组装的动力学；（iv）A亚基在体外不能与完全组装好的B五聚体结合；（v）A亚基在体内使B亚基五聚化速率加快约3倍，这意味着A亚基在全毒素组装途径中起协调作用。最后这一发现可能具有普遍意义，因为它为多亚基蛋白质组装过程中优先排除或促进某些中间体提供了一种机制。
